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www.telstar-lifesciences.com
HexaScreen®
Biotechnological Screening in Animal Cell Culture
Equipment to reduce required time &
costs for bioprocess development
EXPOQUIMIA 2011
www.telstar-lifesciences.com
++
The Team
•Manufacturing
•Marketing & Sales
•Internationalization
•Cell Culture
•Bioreactors
• Measurement Systems
•Biocompatible materials
+
www.telstar-lifesciences.com
Increasing importance of animal cell culture.
www.telstar-lifesciences.com
The potential of animal cell culture arises from the capability of this type of
cells to carry out complex post-translational modifications providing proteins
with the required biological activity to be used for therapeutical and
diagnostic applications
CHO (Chineese Hamster Ovary)
NSO (mouse mieloma)
BHK (Baby Hamster Kidney)
HEK (Human Embryo Kidney)
PER.C6 (Human Retinal Cells)
MDCK (Madin Darby Canine Kidney)
Sf9 (insect cells, Spodoptera frugiperda)
Animal Cell Culture Bioprocesses
www.telstar-lifesciences.com
Bioprocess Development: Animal Cell Culture
”The optimization of the bioprocess is specially important for animal cell culture”
– Less Cellular Concentration than Bacterium and Yeast:
• Bacterium and Yeast cultures: [X]f ≈ 109 cells/ml
• Animal cell cultures: [X]f ≈ 106 cells/ml
– More expensive & complex process:
• Culture media:
– Bacterium and Yeast cell culture: Simple undefined mediums
containing only salts (usually NaCl) and carbon & nitrogen sources
(usually within yeast extract and tryptone).
– Animal cell culture: Rich & complex mediums containing salts,
carbon & nitrogen sources, but also complements (AA, vitamins,
trace elements…) and serums (FCB, FBS…).
• Easier to be contaminated:
– Bacteria, yeast, mycoplasma or cross contamination.
– Specific challenges for animal cell culture: slow growth and specific
production rates, cell sensitivity (shear stress, nutrient limitation,
metabolite accumulation..).
If Ps=ct, we have 1000 times more product
with bacterium or yeast than with animal cells
www.hexascreen.com
from F. Wurm. Nat. Biotechnol., vol. 22:1393-1398 (2004)
Bioprocess Development: Main Steps
Genetic Engineering Culture Conditions Operational Conditions
Best Clones:
Specific Productivity
Best Conditions:
[X]
Growth rate
Optimal Process:
[X]
Death times
www.telstar-lifesciences.com
PROCESS
OPTIMIZATIONClonal
selection
Cell
line
Operation
strategy
Process
control
Purification
process
Scale-up
GMP
production
Culture
medium
Fisico-chemical
parameters
PROCESS
OPTIMIZATION
PROCESS
OPTIMIZATIONClonal
selection
Clonal
selection
Cell
line
Cell
line
Operation
strategy
Operation
strategy
Process
control
Process
control
Purification
process
Purification
process
Scale-up
GMP
production
GMP
production
Culture
medium
Culture
medium
Fisico-chemical
parameters
Fisico-chemical
parameters
Multiple aspects need to be optimized to establish optimal
and profitable production processes
High cell concentration, high specific productivity, maintain product quality
www.telstar-lifesciences.com
Bioprocess Development: Phases
• PHASE I: Initial screening
1. Cell line selection (bacterium, yeast, animal cell…)
2. Cell modification (genetic engineering)
3. Clonal activity selection
Select which cells are producing the protein target
4. Protein target characterization:
Measurement of the protein activity
• PHASE II: Advanced screening
1. Clonal growth selection (select which cells grow faster…)
2. Culture medium composition: serum, glutamine, glucose…
3. Initial cell concentration.
4. Effects of stirring, [O2] and other culture conditions
5. Needs of the cell culture adaptation (ex: from adherent to suspension)
• PHASE III: Lab scale production
1. Scale-up the advanced screening optimum conditions
2. Purification process
3. Type of Bioreactor (Batch, Fed-Batch, Perfusion)
• PHASE IV: Pilot Plant and Industrial scale production
1. Scale-up the lab scale optimum production conditions
7/8
clones
1/2
clones
IMPORTANT!!!
Check the
activity of the
protein in all
phases
www.telstar-lifesciences.com
– Reproducibility problems when scaling up from usual
screening phases into lab or pilot & production plant
scales due to:
• Homogeneity: Non agitated systems can produce cells or nutrient
accumulations giving not representative and impossible to repeat
experiments.
• [O2] limitations: Can produce a decrease of the cell culture growth
velocity, metabolic differences or stopping the cellular cycle leading to an
unreal productivity determination (higher or lower).
– Lack of probes: few knowledge of the main cellular
growth parameters : cell concentration, pH & pO2.
– Lack of automatization: human manipulation is highly
needed.
– Current screening agitated systems (spinner flasks)
characteristics include high volumes and post-
experiment treatments.
Screening Bioreactors: Differences
Cost and time
issues
Reproducibility
problems
www.hexascreen.com
Vessel volume (ml)
0,1 1 10 100 1000+
T-flask
Spinner flask
Bioreactors
Multiwell plates
Advanced Screening
Usual scale-up procedure
New automated & controlled
screening platform
Initial Screening Lab ScaleInitial Screening
Screening : The HexaScreen Alternative
www.telstar-lifesciences.com
Bioprocess Development: Phases
Bioprocess development for a biotechnology product
requires a number of steps
Scale-up Methodology: Laboratory to Pilot to
Industrial
Since Biotechnological Processes are not
completely known, the transition from bench to
final volume is done step by step.
Initial
Screening
Pilot Plant
Advanced
Screening
Production Plant
Lab scale
HEXABATCH
www.hexascreen.com
– Simultaneous multiple experiment capability.
– Low minibioreactor volumes (10-15 mL), but not too low
to allow cell culture similarities with lab scale
bioreactors.
– Agitation requirement to allow system homogeneity, but
without upsetting cell viability.
– Parts in contact with cells must be manufactured with
single use biocompatible plastic (no contamination,…).
– Cell growth (optical density), pH and dissolved oxygen
monitoring via non invasive probes.
Screening: HexaBatch®Design Criteria
www.telstar-lifesciences.com
HexaScreen®: HexaBatch Features
Initial/Advanced
Screening
HexaBatch
Pilot Plant
/ Industry
Equipment used &
environment
Stationary Culture
Systems: Heterogeneous
Stirred & O2 fed Culture
System to allow
homogeneity
Stirred Culture
Systems:
Homogeneous
Methodology Multiple experiments
Multiple experiments run
simultaneously to
decrease the time
required for the
screening phase
Unique cell
culture process
Vessel size Micro liters and milliliters
10-15 ml (bench top
scale)
From 2 to
thousands of
liters
Process control
Discontinuous and
manual
pH, pO2 and OD (cell
concentration) on-line
monitoring
Continuous
and monitored
Asepsis Difficult / uncontrolled
Disposable bioreactor
made of sterile
biocompatible plastic
material
Necessary
www.telstar-lifesciences.com
– Single use Minibioreactors’ Plate. Previously sterilized
inside a plastic bag, includes 6 individual vessels equipped
with gas filters, one septum for inoculation, miniaturized
ports for probe’s allocation and a magnetic actuator for
stirring.
– Workstation. Contains the chamber where the
minibioreactors’s plate remains during its culture, while
maintaining optimal agitation & temperature (common),
sterility, providing individual aeration and acquiring pH, DO
and OD data. WorkStation is controlled via software.
HexaScreen®: HexaBatch Elements
HexaBatch version consists in two differentiated parts,
the 6-minibioreactors’ plate and the workstation with a
computer/software .
www.telstar-lifesciences.com
HexaBatch: Minibioreactor plate characteristics
5
2
1
43
1-. Optical port for OD and pH measurements.
2-. Gas filters (inlet and outlet).
3-. Optical port for DO measurements via
fluorescence.
4-. Septum for cell inoculation.
5-. Low shear magnetic pendular
agitation (optimal for animal cell).
6-. Thermostated general bath.
7-. Vessel liquid volume: 10-15 ml.
8-. Biocompatible and disposable plastic,
sterile provided. Possible plasma treatment
to promote cell adherence / non-
adherence.
7 6
8
www.hexascreen.com
HexaBatch: Minibioreactors Inoculation
www.hexascreen.com
HexaBatch: Minibioreactors to Workstation
www.telstar-lifesciences.com
HexaBatch: Variables, Controls & Monitoring
Variable Control Monitoring Information
Aeration Time Controlled No No
Agitation Set-point controlled No No
Vessel
Temperature
Set-point controlled Monitored System functionality
Gas & Filters
Temperature
Set-point controlled Monitored
Filters functionality –
Avoids evaporation
Optical Density Free evolution
Monitored and
correlated
Cell density information,
related to total cells
Dissolved
Oxygen
Free evolution
(constant aeration)
Monitored
Oxygen concentration,
related to alive cells
pH Free evolution Monitored
Cell activity information,
related to alive cells
www.hexascreen.com
HexaBatch: Software Specifications
General specifications
• Automatic processes:
– Workstation’s configuration
– Thermal stabilization
– Oxygen calibration
– Optical calibration
– Data acquisition (minibioreactors’ behaviour monitoring graphs)
• Additional tasks:
– Create new or edit already existing experiment set-ups
– Create user defined graphs
– Create reports
– Export reports to EXCEL, PDF and HTML files
HexaScreen®’s control & acquisition program runs on a Windows
platform and will guide the user, as a wizard, through the processes of
workstation’s configuration, calibration and data acquisition.
www.hexascreen.com
HexaBatch: Advantages & Benefits
Features Advantages Benefits
Specially designed for animal cell
culture.
- Stirring, gas exchange and oxygen supply are
specially designed for handling animal cell
culture.
- Focused.
- Suitable for both suspension and
adherent cultures.
Benchtop Scale:
Small Volumes from 10 to 15 ml”.
- Reducing working volume means less
development costs in medium, cell culture,
enzymes…
- Lower operation cost.
Parallel bioreactor system:
“6 multiple parallel experiments
for device”.
- Less time required to achieve final results
- Reproducible results: statistic data can be
obtained from replicated experiments.
- Faster time to market.
- Save time.
- Faster product development cycles.
Automated on-line
measurements.
- Real-time kinetic information:
cell concentration (OD), DO & pH.
- No off-line control processes required.
- No maintenance required during experiment.
- Precision & control over all
experimentation.
- Lower labour and time-consuming in
order to invest in other valuable
tasks.
Single use.
- No post-experiments treatments.
- Avoid possible cross-contaminations.
- Save money & time.
- Easier experiment validations.
Easy to use. - No expert personal required. - Less effort required.
Convenient. - Easier scale up to lab-size reactors. - Optimized culture parameters.
PC controlled.
- Save automatically the data acquired from all
the experiments done giving a comprehensive
documentation.
- Control on all acquisition
experiment data.
- Possibility to do final reports easier.
www.hexascreen.com
HexaBatch: Applications & Cell Lines Cultured
Applications Examples
Cellular screening.
- Adherence and suspension lines.
- Clone selection.
Cellular characterization.
- Growth parameters measurements (cell
density and cell activity).
Cellular adaptation. - Adherent to suspension.
Medium definition and
optimization.
- Commercial medium comparison.
- Medium’s components definition.
Cellular tests.
- New drugs tests.
- Toxicity tests.
- Apoptosis tests.
Process optimization.
- Initial cellular concentration, culture
conditions…
Cell Lines Cultured
Suspension:
- Hybridoma.
- Genetically modified hybridoma.
- CHO cells (adapted).
- HEK (adapted)
Adherent:
- Vero cells.
- Ovine Mesenchymal Stem Cells.
- CHO.
- HEK.
www.hexascreen.com
 Fast setup.
 No need for Cleaning.
 No risk of cross-contamination.
 Minimizes utility requirements.
 Minimizes validation.
 Minimizes space floor.
 Minimizes labor.
 Minimizes engineering design.
 Reduces COGS.
 Minimizes maintenance.
 Environmentally friendly:
 Use for generate electrical power by incineration.
 Estimated savings in WFI at over 80%.
 Estimated 72% saving in electricity compared to conventional
manufacturing facility.
Advantages of single-use technology
www.hexascreen.com
Advantages of single-use technology
www.telstar-lifesciences.com
• Systems to evaluate:
• HexaBatch
• 1L glass Bioreactor
• T-flasks for suspension cell cultures
• T-flasks for adherent cell cultures
Systems comparison for screening
On-line measurements
Off-line measurements
www.hexascreen.com
• Case Study parameters:
• Number of conditions: 2
• Number of repetitions: 3
• Total number of cultures: 6
• Batch culture time: 3 days
• Cell line: CHO
• Culture media price (CDCHO): 103,52 €/L
• Qualified personnel costs: 30 €/hour
• It is assumed that there is only a one-liter bioreactor, so steps of the
experiment are performed sequentially.
• For off-line measurements (T-Flasks), only one sample is taken each
day, with a total of 3 cell concentration measurements during culture.
(no metabolites concentration measured)
HexaBatch Case Study: Conditions
www.hexascreen.com
1
7
1 2
18
3
18
3
3
0
5
10
15
20
25
30
35
40
45
1 HexaBatch 1L Bioreactor T-flasks T-flasks (adherent cell)
Experimentaltime(days)
4 days
43 days
4 days
5 days
699 Total personnel cost (€)
Total bioreactor operation cost (€)
Total culture medium cost (€)
Fins a
5040
totals
Fin
19
tot
Inoculum Scale-up time
Bioreactor set-up time
(Set-up, sterilization, CIP, calibration, post-exp. cleaning)
Culture time
36 days
HexaBatch Case Study: Time Analysis
www.hexascreen.com
HexaBatch Case Study: Costs Analysis
699
21
21
12
978
1800
0
500
1000
1500
2000
2500
1 HexaBatch 1L Bioreactor T-flasks T-flasks (adherent cell)
Costedeunexperimento(€)
699
151
12
22,5
150
219 74
Totalexperimentalcost(€)
Total personnel cost (€)
Total bioreactor operation cost (€)
Total culture medium cost (€)
Fins a
5040
totals
Fins a
1960
totals699
151
12
22,5
150
219 74
Totalexperimentalcost(€)
Total personnel cost (€)
Total bioreactor operation cost (€)
Total culture medium cost (€)
Fins a
5040
totals
Fins a
1960
totals
699 Total personnel cost (€)
Total bioreactor operation cost (€)
Total culture medium cost (€)
Fins a
5040
totals
Fi
1
to
Personnel costs
(staff hours x costs/hour)
Lab material costs
(Single-use plate costs + T-flask costs for scale-up)
Culture medium costs
183 €
2.520 €
161 €
507 €
699
151
12
22,5
150
219 74
Totalexperimentalcost(€)
Total personnel cost (€)
Total bioreactor operation cost (€)
Total culture medium cost (€)
Fins a
5040
totals
Fins a
1960
totals
699
151
12
22,5
150
219 74
Totalexperimentalcost(€)
Total personnel cost (€)
Total bioreactor operation cost (€)
Total culture medium cost (€)
Fins a
5040
totals
Fins a
1960
totals
Experimentcosts(€)
www.hexascreen.com
HexaBatch Case Study: Manipulation Times
33 hours
45 min
60 hours
5 hours
0
10
20
30
40
50
60
1 HexaBatch 1L Bioreactor T-flasks T-flasks (adherent cell)
QualifiedPersonnelManipulationTime(hours)
www.hexascreen.com
HexaBatch Results: On-line Optical Density
Total cells
, tdup
Final cell concentration
Off-line
Measurements
(T-FLASK)
- Hexabatch gives one on-line OD measurement every five minutes vs
just one/two per day in the case of T-flasks.
www.hexascreen.com
HexaBatch Results: On-line pH Measurements
Gives
information
about cell
activity
Faster response than OD
(no death cells interaction)
, tdup (indirect)
www.hexascreen.com
HexaBatch Results: On-line Dissolved Oxygen
Optimal Range
www.telstar-lifesciences.com
HexaBatch System
=
1L bioreactor specifications
+
T-Flasks cheap and fast experimentation
• Technological advantages from a bioreactor
• System’s homogeneity: stirring and aeration
• On-line process monitoring (pH, DO, OD)
• Easy maintenance of asepsis
• T-flask experiment price, speed and flexibility
• Working volume at ml scale
• Multiple experiment capability
• Minimal needs on qualified personnel
Conclusion: HexaBatch system
www.hexascreen.com
HexaScreen Applications: Index
1. Pharmacokinetic Tests.
2. Clone Comparison.
3. Media Comparison.
4. Inoculum Concentration.
5. Adaption to Suspension Cultures.
www.hexascreen.com
Drug Functional Tests: Pharmacokinetics
Antibiotic effect in cell cultures
Functional Tests
Activity profiles
More advantages to perform functional tests
in cell cultures than in animals:
- Faster results
- Ethical issues
www.hexascreen.com
Advanced Screening: Clone Comparison
Chose the best clone in terms of:
- Cell growth rates.
- Cell activity.
- Productivity at different sample times activity
activity
Growth
rates
www.hexascreen.com
Advanced Screening: Media Comparison
• Best growth medium
• Time of action:
- Fed-Batch start point.
- Infection.
- Product recovery.
Medium
component
depletion
www.hexascreen.com
Cell concentration profiles
obtained from pH profiles
(related to cell activity)
Advanced Screening: Inoculum Concentration
www.hexascreen.com
Advanced Screening: Suspension Adaption
Control over adaption process
Cell concentration control along passages
www.hexascreen.com
JAVIER AMAYRA: jamayra@hexascreen.com General Manager
GRACIAS POR SU ATENCIÓN:
HexaScreen Culture Technologies S.L.
Edifici Eureka, P1M1.2
Parc de Recerca de la Universidad Autónoma de Barcelona (UAB)
08193 Cerdanyola del Vallès (Barcelona)

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Expoquimia 2011: Forum Biotech - Javier Amaya

  • 1. www.telstar-lifesciences.com HexaScreen® Biotechnological Screening in Animal Cell Culture Equipment to reduce required time & costs for bioprocess development EXPOQUIMIA 2011
  • 2. www.telstar-lifesciences.com ++ The Team •Manufacturing •Marketing & Sales •Internationalization •Cell Culture •Bioreactors • Measurement Systems •Biocompatible materials +
  • 4. www.telstar-lifesciences.com The potential of animal cell culture arises from the capability of this type of cells to carry out complex post-translational modifications providing proteins with the required biological activity to be used for therapeutical and diagnostic applications CHO (Chineese Hamster Ovary) NSO (mouse mieloma) BHK (Baby Hamster Kidney) HEK (Human Embryo Kidney) PER.C6 (Human Retinal Cells) MDCK (Madin Darby Canine Kidney) Sf9 (insect cells, Spodoptera frugiperda) Animal Cell Culture Bioprocesses
  • 5. www.telstar-lifesciences.com Bioprocess Development: Animal Cell Culture ”The optimization of the bioprocess is specially important for animal cell culture” – Less Cellular Concentration than Bacterium and Yeast: • Bacterium and Yeast cultures: [X]f ≈ 109 cells/ml • Animal cell cultures: [X]f ≈ 106 cells/ml – More expensive & complex process: • Culture media: – Bacterium and Yeast cell culture: Simple undefined mediums containing only salts (usually NaCl) and carbon & nitrogen sources (usually within yeast extract and tryptone). – Animal cell culture: Rich & complex mediums containing salts, carbon & nitrogen sources, but also complements (AA, vitamins, trace elements…) and serums (FCB, FBS…). • Easier to be contaminated: – Bacteria, yeast, mycoplasma or cross contamination. – Specific challenges for animal cell culture: slow growth and specific production rates, cell sensitivity (shear stress, nutrient limitation, metabolite accumulation..). If Ps=ct, we have 1000 times more product with bacterium or yeast than with animal cells
  • 6. www.hexascreen.com from F. Wurm. Nat. Biotechnol., vol. 22:1393-1398 (2004) Bioprocess Development: Main Steps Genetic Engineering Culture Conditions Operational Conditions Best Clones: Specific Productivity Best Conditions: [X] Growth rate Optimal Process: [X] Death times
  • 8. www.telstar-lifesciences.com Bioprocess Development: Phases • PHASE I: Initial screening 1. Cell line selection (bacterium, yeast, animal cell…) 2. Cell modification (genetic engineering) 3. Clonal activity selection Select which cells are producing the protein target 4. Protein target characterization: Measurement of the protein activity • PHASE II: Advanced screening 1. Clonal growth selection (select which cells grow faster…) 2. Culture medium composition: serum, glutamine, glucose… 3. Initial cell concentration. 4. Effects of stirring, [O2] and other culture conditions 5. Needs of the cell culture adaptation (ex: from adherent to suspension) • PHASE III: Lab scale production 1. Scale-up the advanced screening optimum conditions 2. Purification process 3. Type of Bioreactor (Batch, Fed-Batch, Perfusion) • PHASE IV: Pilot Plant and Industrial scale production 1. Scale-up the lab scale optimum production conditions 7/8 clones 1/2 clones IMPORTANT!!! Check the activity of the protein in all phases
  • 9. www.telstar-lifesciences.com – Reproducibility problems when scaling up from usual screening phases into lab or pilot & production plant scales due to: • Homogeneity: Non agitated systems can produce cells or nutrient accumulations giving not representative and impossible to repeat experiments. • [O2] limitations: Can produce a decrease of the cell culture growth velocity, metabolic differences or stopping the cellular cycle leading to an unreal productivity determination (higher or lower). – Lack of probes: few knowledge of the main cellular growth parameters : cell concentration, pH & pO2. – Lack of automatization: human manipulation is highly needed. – Current screening agitated systems (spinner flasks) characteristics include high volumes and post- experiment treatments. Screening Bioreactors: Differences Cost and time issues Reproducibility problems
  • 10. www.hexascreen.com Vessel volume (ml) 0,1 1 10 100 1000+ T-flask Spinner flask Bioreactors Multiwell plates Advanced Screening Usual scale-up procedure New automated & controlled screening platform Initial Screening Lab ScaleInitial Screening Screening : The HexaScreen Alternative
  • 11. www.telstar-lifesciences.com Bioprocess Development: Phases Bioprocess development for a biotechnology product requires a number of steps Scale-up Methodology: Laboratory to Pilot to Industrial Since Biotechnological Processes are not completely known, the transition from bench to final volume is done step by step. Initial Screening Pilot Plant Advanced Screening Production Plant Lab scale HEXABATCH
  • 12. www.hexascreen.com – Simultaneous multiple experiment capability. – Low minibioreactor volumes (10-15 mL), but not too low to allow cell culture similarities with lab scale bioreactors. – Agitation requirement to allow system homogeneity, but without upsetting cell viability. – Parts in contact with cells must be manufactured with single use biocompatible plastic (no contamination,…). – Cell growth (optical density), pH and dissolved oxygen monitoring via non invasive probes. Screening: HexaBatch®Design Criteria
  • 13. www.telstar-lifesciences.com HexaScreen®: HexaBatch Features Initial/Advanced Screening HexaBatch Pilot Plant / Industry Equipment used & environment Stationary Culture Systems: Heterogeneous Stirred & O2 fed Culture System to allow homogeneity Stirred Culture Systems: Homogeneous Methodology Multiple experiments Multiple experiments run simultaneously to decrease the time required for the screening phase Unique cell culture process Vessel size Micro liters and milliliters 10-15 ml (bench top scale) From 2 to thousands of liters Process control Discontinuous and manual pH, pO2 and OD (cell concentration) on-line monitoring Continuous and monitored Asepsis Difficult / uncontrolled Disposable bioreactor made of sterile biocompatible plastic material Necessary
  • 14. www.telstar-lifesciences.com – Single use Minibioreactors’ Plate. Previously sterilized inside a plastic bag, includes 6 individual vessels equipped with gas filters, one septum for inoculation, miniaturized ports for probe’s allocation and a magnetic actuator for stirring. – Workstation. Contains the chamber where the minibioreactors’s plate remains during its culture, while maintaining optimal agitation & temperature (common), sterility, providing individual aeration and acquiring pH, DO and OD data. WorkStation is controlled via software. HexaScreen®: HexaBatch Elements HexaBatch version consists in two differentiated parts, the 6-minibioreactors’ plate and the workstation with a computer/software .
  • 15. www.telstar-lifesciences.com HexaBatch: Minibioreactor plate characteristics 5 2 1 43 1-. Optical port for OD and pH measurements. 2-. Gas filters (inlet and outlet). 3-. Optical port for DO measurements via fluorescence. 4-. Septum for cell inoculation. 5-. Low shear magnetic pendular agitation (optimal for animal cell). 6-. Thermostated general bath. 7-. Vessel liquid volume: 10-15 ml. 8-. Biocompatible and disposable plastic, sterile provided. Possible plasma treatment to promote cell adherence / non- adherence. 7 6 8
  • 18. www.telstar-lifesciences.com HexaBatch: Variables, Controls & Monitoring Variable Control Monitoring Information Aeration Time Controlled No No Agitation Set-point controlled No No Vessel Temperature Set-point controlled Monitored System functionality Gas & Filters Temperature Set-point controlled Monitored Filters functionality – Avoids evaporation Optical Density Free evolution Monitored and correlated Cell density information, related to total cells Dissolved Oxygen Free evolution (constant aeration) Monitored Oxygen concentration, related to alive cells pH Free evolution Monitored Cell activity information, related to alive cells
  • 19. www.hexascreen.com HexaBatch: Software Specifications General specifications • Automatic processes: – Workstation’s configuration – Thermal stabilization – Oxygen calibration – Optical calibration – Data acquisition (minibioreactors’ behaviour monitoring graphs) • Additional tasks: – Create new or edit already existing experiment set-ups – Create user defined graphs – Create reports – Export reports to EXCEL, PDF and HTML files HexaScreen®’s control & acquisition program runs on a Windows platform and will guide the user, as a wizard, through the processes of workstation’s configuration, calibration and data acquisition.
  • 20. www.hexascreen.com HexaBatch: Advantages & Benefits Features Advantages Benefits Specially designed for animal cell culture. - Stirring, gas exchange and oxygen supply are specially designed for handling animal cell culture. - Focused. - Suitable for both suspension and adherent cultures. Benchtop Scale: Small Volumes from 10 to 15 ml”. - Reducing working volume means less development costs in medium, cell culture, enzymes… - Lower operation cost. Parallel bioreactor system: “6 multiple parallel experiments for device”. - Less time required to achieve final results - Reproducible results: statistic data can be obtained from replicated experiments. - Faster time to market. - Save time. - Faster product development cycles. Automated on-line measurements. - Real-time kinetic information: cell concentration (OD), DO & pH. - No off-line control processes required. - No maintenance required during experiment. - Precision & control over all experimentation. - Lower labour and time-consuming in order to invest in other valuable tasks. Single use. - No post-experiments treatments. - Avoid possible cross-contaminations. - Save money & time. - Easier experiment validations. Easy to use. - No expert personal required. - Less effort required. Convenient. - Easier scale up to lab-size reactors. - Optimized culture parameters. PC controlled. - Save automatically the data acquired from all the experiments done giving a comprehensive documentation. - Control on all acquisition experiment data. - Possibility to do final reports easier.
  • 21. www.hexascreen.com HexaBatch: Applications & Cell Lines Cultured Applications Examples Cellular screening. - Adherence and suspension lines. - Clone selection. Cellular characterization. - Growth parameters measurements (cell density and cell activity). Cellular adaptation. - Adherent to suspension. Medium definition and optimization. - Commercial medium comparison. - Medium’s components definition. Cellular tests. - New drugs tests. - Toxicity tests. - Apoptosis tests. Process optimization. - Initial cellular concentration, culture conditions… Cell Lines Cultured Suspension: - Hybridoma. - Genetically modified hybridoma. - CHO cells (adapted). - HEK (adapted) Adherent: - Vero cells. - Ovine Mesenchymal Stem Cells. - CHO. - HEK.
  • 22. www.hexascreen.com  Fast setup.  No need for Cleaning.  No risk of cross-contamination.  Minimizes utility requirements.  Minimizes validation.  Minimizes space floor.  Minimizes labor.  Minimizes engineering design.  Reduces COGS.  Minimizes maintenance.  Environmentally friendly:  Use for generate electrical power by incineration.  Estimated savings in WFI at over 80%.  Estimated 72% saving in electricity compared to conventional manufacturing facility. Advantages of single-use technology
  • 24. www.telstar-lifesciences.com • Systems to evaluate: • HexaBatch • 1L glass Bioreactor • T-flasks for suspension cell cultures • T-flasks for adherent cell cultures Systems comparison for screening On-line measurements Off-line measurements
  • 25. www.hexascreen.com • Case Study parameters: • Number of conditions: 2 • Number of repetitions: 3 • Total number of cultures: 6 • Batch culture time: 3 days • Cell line: CHO • Culture media price (CDCHO): 103,52 €/L • Qualified personnel costs: 30 €/hour • It is assumed that there is only a one-liter bioreactor, so steps of the experiment are performed sequentially. • For off-line measurements (T-Flasks), only one sample is taken each day, with a total of 3 cell concentration measurements during culture. (no metabolites concentration measured) HexaBatch Case Study: Conditions
  • 26. www.hexascreen.com 1 7 1 2 18 3 18 3 3 0 5 10 15 20 25 30 35 40 45 1 HexaBatch 1L Bioreactor T-flasks T-flasks (adherent cell) Experimentaltime(days) 4 days 43 days 4 days 5 days 699 Total personnel cost (€) Total bioreactor operation cost (€) Total culture medium cost (€) Fins a 5040 totals Fin 19 tot Inoculum Scale-up time Bioreactor set-up time (Set-up, sterilization, CIP, calibration, post-exp. cleaning) Culture time 36 days HexaBatch Case Study: Time Analysis
  • 27. www.hexascreen.com HexaBatch Case Study: Costs Analysis 699 21 21 12 978 1800 0 500 1000 1500 2000 2500 1 HexaBatch 1L Bioreactor T-flasks T-flasks (adherent cell) Costedeunexperimento(€) 699 151 12 22,5 150 219 74 Totalexperimentalcost(€) Total personnel cost (€) Total bioreactor operation cost (€) Total culture medium cost (€) Fins a 5040 totals Fins a 1960 totals699 151 12 22,5 150 219 74 Totalexperimentalcost(€) Total personnel cost (€) Total bioreactor operation cost (€) Total culture medium cost (€) Fins a 5040 totals Fins a 1960 totals 699 Total personnel cost (€) Total bioreactor operation cost (€) Total culture medium cost (€) Fins a 5040 totals Fi 1 to Personnel costs (staff hours x costs/hour) Lab material costs (Single-use plate costs + T-flask costs for scale-up) Culture medium costs 183 € 2.520 € 161 € 507 € 699 151 12 22,5 150 219 74 Totalexperimentalcost(€) Total personnel cost (€) Total bioreactor operation cost (€) Total culture medium cost (€) Fins a 5040 totals Fins a 1960 totals 699 151 12 22,5 150 219 74 Totalexperimentalcost(€) Total personnel cost (€) Total bioreactor operation cost (€) Total culture medium cost (€) Fins a 5040 totals Fins a 1960 totals Experimentcosts(€)
  • 28. www.hexascreen.com HexaBatch Case Study: Manipulation Times 33 hours 45 min 60 hours 5 hours 0 10 20 30 40 50 60 1 HexaBatch 1L Bioreactor T-flasks T-flasks (adherent cell) QualifiedPersonnelManipulationTime(hours)
  • 29. www.hexascreen.com HexaBatch Results: On-line Optical Density Total cells , tdup Final cell concentration Off-line Measurements (T-FLASK) - Hexabatch gives one on-line OD measurement every five minutes vs just one/two per day in the case of T-flasks.
  • 30. www.hexascreen.com HexaBatch Results: On-line pH Measurements Gives information about cell activity Faster response than OD (no death cells interaction) , tdup (indirect)
  • 31. www.hexascreen.com HexaBatch Results: On-line Dissolved Oxygen Optimal Range
  • 32. www.telstar-lifesciences.com HexaBatch System = 1L bioreactor specifications + T-Flasks cheap and fast experimentation • Technological advantages from a bioreactor • System’s homogeneity: stirring and aeration • On-line process monitoring (pH, DO, OD) • Easy maintenance of asepsis • T-flask experiment price, speed and flexibility • Working volume at ml scale • Multiple experiment capability • Minimal needs on qualified personnel Conclusion: HexaBatch system
  • 33. www.hexascreen.com HexaScreen Applications: Index 1. Pharmacokinetic Tests. 2. Clone Comparison. 3. Media Comparison. 4. Inoculum Concentration. 5. Adaption to Suspension Cultures.
  • 34. www.hexascreen.com Drug Functional Tests: Pharmacokinetics Antibiotic effect in cell cultures Functional Tests Activity profiles More advantages to perform functional tests in cell cultures than in animals: - Faster results - Ethical issues
  • 35. www.hexascreen.com Advanced Screening: Clone Comparison Chose the best clone in terms of: - Cell growth rates. - Cell activity. - Productivity at different sample times activity activity Growth rates
  • 36. www.hexascreen.com Advanced Screening: Media Comparison • Best growth medium • Time of action: - Fed-Batch start point. - Infection. - Product recovery. Medium component depletion
  • 37. www.hexascreen.com Cell concentration profiles obtained from pH profiles (related to cell activity) Advanced Screening: Inoculum Concentration
  • 38. www.hexascreen.com Advanced Screening: Suspension Adaption Control over adaption process Cell concentration control along passages
  • 39. www.hexascreen.com JAVIER AMAYRA: jamayra@hexascreen.com General Manager GRACIAS POR SU ATENCIÓN: HexaScreen Culture Technologies S.L. Edifici Eureka, P1M1.2 Parc de Recerca de la Universidad Autónoma de Barcelona (UAB) 08193 Cerdanyola del Vallès (Barcelona)